The objectives of the project are to understand the enzymatic and genetic basis for the extraordinary nutritional versatility of the opportunistic pathogen Pseudomonas cepacia--including its unusual capacity to utilize beta-lactam antibiotics as sole carbon and energy source. We have defined the roles of two beta-lactamase iso-enzymes in dissimilation of penicillin G and cephalosporin C, and now are proceeding to identify other enzymes involved in this process. We are continuing our characterization of unusual pleiotropic mutants that wer originally selected on the basis of their lysine requirement and later found unable to utilize penicillin G, ribitol, orlysine as carbon source. These strains were also deficient in beta-lactamase. Recently we discovered that the mutants have an alteration in the 95 Mdal plasmid derived from the wild type strain 249. Restriction enzyme analysis of plasmid DNA from the wild type and mutant strains suggests that the alteration may be reorientation of an invertible region on the plasmid. A major goal of the research in the coming year will be to further define the plasmid alteration in the strains we have on hand and to obtain and examine new mutants directly selected for failure to utilize penicillin, ribitol, or lysine. Another aim of the work is development of systems of genetic analysis which will facilitate studies of gene expression in these bacteria. Towards this end we are continuing our efforts to develop efficient conjugation and transduction systems utilizing P. cepacia strains which harbor the R factors RPl and R68.45 and a newly isolated phage, CP5, which promises to be suitable for transductional analysis of P. cepacia.